Inland Reef Aquaria Salt Study, Part I

Background

In March, 2003, Dr. Ronald Shimek
published the results of a bio-assay study in which he concluded
that two of the most popular salt mixes in the aquarium industry,
Instant Ocean and Coralife, were toxic to sea life
(1). The study further concluded that two
other less common salt mixes, Bio-Sea MarineMix and Crystal
Sea Bio-Assay Formula, fared significantly better – comparable
to natural sea water (2). Dr. Shimek indicated that the most
likely cause of this toxicity was the level of heavy metals
present in the salt mixes (3).

Like most aquarists, we were immediately concerned over these
results. As the title of Dr. Shimek’s work states, toxic
metals in our salt mixes are indeed a bad beginning for a reef
aquarium. Further inspection of the study revealed several areas
in which further testing and study might be of value.

First, the data on heavy metal concentrations for these four
salt mixes had come from two very different sources: Instant
Ocean and Coralife from a four year old independent analysis
(Atkinson & Bingman, 1999) and the other two from the
manufacturer. Dr. Shimek does express some concern over this
(4), but concludes that the
manufacturer’s data is not suspect (5). This was an interesting conclusion in
light of the work that Dr. Shimek and we had done on Two
Little Fishies Combisan two years prior (Inland Reef and Ron
Shimek 2001). In that case, the data printed on the bottle,
from a very well-respected manufacturer, was found by Dr.
Shimek to be completely unreliable. In the controversy that
followed, Inland Reef conducted an independent analysis and
confirmed Dr. Shimek’s data, eventually resulting in the
manufacturer removing the printed analysis from their
bottle.

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Second, if obviously high toxic metals levels in Instant Ocean
and Coralife were the root cause of the poor bio-assay
performance in Dr. Shimek’s study, we reasoned that
conducting an analysis on all four salts, handled and testing in
an identical manner, might reveal the causative element or
elements.

Finally, we had it good authority that the chemical formulas
for many salt mixes had been updated since Atkinson and
Bingman’s 1999 study. This valuable data was badly in need of
being updated using today’s salt mix formulations.

We therefore resolved to conduct an independent elemental
analysis of the salt mixes. After discussions on Reefs.org, the
scope of the work grew to cover all of the popular salt mixes
currently used in the hobby, not just those in Dr. Shimek’s
study. The high cost of performing so many elemental analyses led
Inland Reef to seek donations to assist with the total costs. The
vast majority of funding for this study was provided by members
of Reefs.org and several auctions of donated merchandise
conducted by Reefs.org. There were also contributions from the
members of ReefCentral.com and a matching donation from the
ReefCentral.com staff. The remainder of the cost was covered by
Inland Reef Aquaria directly.

Unfortunately, we made the decision to close Inland Reef
Aquaria and pursue other business interests just as the study was
getting underway. This and several other “real-world”
responsibilities led to repeated delays in the study and the
publication of its results. We apologize sincerely to all of you
who have contributed to this effort and have patiently awaited
the results for far longer than anyone ever anticipated. We hope
you will find it worth the wait!

Procedure

The selection of salts to be tested was based on a series of
polls conducted on the Reefs.org discussion forum. Only a limited
number of tests could be performed with the available funds. In
addition, poll results favored the inclusion of Oceanic’s new
salt mix over Coralife, causing one of the salts in Dr.
Shimek’s study to be dropped from the list. The final list,
in no particular order, is:

AquaMedic Sea Salt (AM)

Oceanic Sea Salt (O)

Omega Sea Marine Salt (OS)

Aqua Craft Bio-Sea MarineMix (BSM)

Marine Enterprises Crystal Sea MarineMix (CSM)

Tropic Marin Sea Salt (TM)

Instant Ocean (IO)

Kent Marine Salt (KM)

Marine Enterprises Crystal Sea Bio-Assay Formula (CSB)

The abbreviations indicated here are used on the graphs and
tables to be found below.

Several bags (or boxes) of each salt were obtained over a 10
month period. All salts were either purchased from wholesale
suppliers or retail stores, or obtained directly from the
manufacturer at trade shows. In no case was any manufacturer
aware that they were providing a sample to be used in this
study.

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Several samples were obtained from each bag, from various
positions within the bag, to minimize the effects of settling. In
a perfect world, several elemental analyses would be performed on
each salt brand, allowing an error range to be determined for
each element tested. Unfortunately, the high cost of these
analyses ruled out performing that many tests – an initial
estimate to perform three tests per salt mix came to over
$15,000. Therefore, the samples from at least three different
bags of the same brand salt were combined to obtain an
“average” sample for each brand. We then made sure that
the testing lab performed each batch in succession with
calibration runs interleaved. We also included a reference sample
of natural sea water and RO/DI water with each run. The raw data
for the NSW and RO/DI samples gave us a very high confidence in
the data for all elements tested. In addition, we are reasonably
certain that the period over which we obtained the sample bags
did not include formula changes by any manufacturer.

Once the samples were taken, each was assigned a random code
letter. All subsequent handling and testing was performed
“in the blind” by someone who did not know the code
letter assignments. Only after the results came back from the lab
were they re-associated with the actual salt mix names.

All labware that came into contact with the salts, including
mixing and measuring containers, sample containers, stirring
rods, sampling spoons, etc., were triple acid-washed
polypropylene to minimize the possibility of any metals
contamination. Separate sets of labware were used for each salt
mix to prevent any cross-contamination.

The salts were mixed with RO/DI water to a concentration of 35
ppt, as measured with a calibrated refractometer. A sample of the
RO/DI water was included in each batch of samples sent to the
lab. The RO/DI water tested below detection limits on all
elements except Zinc, which measured 21 ppb. This value was
subtracted from the Zinc levels reported for each salt
sample.

Yield Test

To test yield, we measured the dry weight of each salt mix
required to produce a given amount of salt water at a
concentration of 35 ppt. To keep moisture accumulation to a
minimum, the length of time each dry sample was exposed to the
air was minimized and all dry samples were handled in an
identical manner. After all the salt solutions were mixed, the
salinity was tested with a calibrated refractometer. The wet
samples were then allowed 24 hours to fully dissolve before
re-testing and, if needed, adjusting the salinity to 35 ppt. The
table below shows the amount of dry sample used, the volume of
salt water produced, and the actual yield from a 50 gallon bag
(or equivalent portion) of each salt:

Salt Mix

Dry Weight (g)

Yield (ml)

ml produced per g

50 gal bag yields

Aqua Medic

21.14

536

25.355

42.26 gal

Oceanic

21.35

526

24.637

42.96 gal

Omega Sea

23.47

571

24.329

43.70 gal

Bio-Sea Marinemix

22.59

580

25.675

46.12 gal

Crystal Sea Marinemix

21.39

534

24.965

43.53 gal

Tropic Marin

21.19

540

25.484

41.74 gal

Instant Ocean

20.70

520

25.121

45.13 gal

Kent

20.67

520

25.157

42.20 gal

Crystal Sea Bio-Assay

20.65

510

24.697

43.00 gal

There is a difference of 4.38 gallons between the highest and
lowest yields (just under a 9% variation). But, clearly, every
salt manufacturer plans for a salinity less than full strength
sea water when advertising the yield of their bags.

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Elemental Testing

Initially, we wanted to perform some basic testing (pH,
Ammonia, Nitrite, and Nitrate) before sending the wet samples to
the lab since the Inductively Coupled Plasma Mass Spectrogram
(ICPMS) testing only gives elemental concentrations. In
preparation, we did some preliminary research on consumer test
kits, as well as performing some basic tests on reference
samples. We came to the unfortunate conclusion that no consumer
test kit on the market today is accurate enough for these
purposes. In some cases, even a basic trend was difficult to
obtain (i.e., higher concentration reference samples sometimes
failed to actually test higher). Preliminary testing was
therefore abandoned and the wet samples were sent off to the
local laboratory. They agreed to test pH prior to ICPMS, but
testing Nitrate or similar compounds would require iron
chromatography or similar tests at a substantially higher cost
per sample.

Silver and Tellurium were below the test’s 5 ppb detection
limit for all salts. Interestingly, Silver levels for Instant
Ocean and Coralife were reported at 248 ppb and 410 ppb
respectively in the 1999 Atkinson and Bingman study.

Gold and Mercury were below their 50 ppb detection limit, and
Iron was below the 500 ppb detection limit for all samples.

The tests for several elements showed little significant
difference between salt mixes. The data for these elements can be
found in the Appendix.

Natural Sea Water (NSW) levels were taken from a variety of
reports and studies. The range of values for NSW element
concentrations varies depending on the study and the area of the
ocean that was sampled. For all elements we list both the lowest
and highest values to establish an acceptable range.

pH

Natural sea water averages pH 8.25. Saltwater aquaria normally
range from 7.8 to 8.5. The measured pH for each mixed salt sample
is shown below. The pH measurement was made 24 hours after the
initial mix and after regular aeration.

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Aluminum (ppb)

This is one of the elements called out in Dr. Shimek’s
study as showing significantly higher toxicity levels for Instant
Ocean and Coralife as compared to Bio-Sea MarineMix and Crystal
Sea Bio-Assay. As can be seen, Bio-Sea MarineMix tested with the
second highest Aluminum concentration and Crystal Sea Bio-Assay
was comparable to Instant Ocean.

Antimony (ppb)

Antimony is a potential heavy metal toxin that was not called
out in Dr. Shimek’s study. The two salts from Marine
Enterprises: Crystal Sea MarineMix and Crystal Sea Bio-Assay,
along with Kent Marine, had significantly higher than NSW levels
(about ten times NSW).

Boron (ppm)

Boron is a not considered a heavy metal. It is a semiconductor
and is lighter than Carbon, Nitrogen, and Oxygen. It is normally
found in seawater as Borate ions. There is a wide variation in
Boron levels in the salt mixes tested, from less than one
thirtieth of NSW values to more than three times NSW.
Interestingly, Crystal Sea Bio-Assay had the second highest Boron
level, Crystal Sea MarineMix, from the same manufacturer, was the
highest, and Instant Ocean was the closest to NSW levels.

Barium (ppb)

Barium is another potential heavy metal contaminant. Both
Bio-Sea MarineMix and Crystal Sea Bio-Assay are similar to NSW.
Instant Ocean tested with the lowest Barium concentration.

Bismuth (ppb)

Bismuth is a potential heavy metal contaminant that is not
called out in Dr. Shimek’s study. All salt mixes except
AquaMedic tested below the 5 ppb detection limit of the test.

Bromine (ppm)

Bromine is a halogen, a non-metallic salt-forming element
related to Chlorine, Fluorine, Iodine, and Astatine. It is
present in sea water mostly as free bromide ions. All salt mixes
have excess levels of Bromine compared to NSW, but it should not
be a toxic contaminant at any of these levels.

Chromium (ppb)

Chromium is another element called out in Dr. Shimek’s
study as showing significantly higher toxicity levels for Instant
Ocean and Coralife as compared to Bio-Sea MarineMix and Crystal
Sea Bio-Assay. Our test results show that, while all salts were
higher than NSW, Instant Ocean was actually the lowest. Bio-Sea
MarineMix was the second highest, and Crystal Sea Bio-Assay had
the third highest Chromium concentration.

Copper (ppb)

Another heavy metal toxin, Copper levels were higher than NSW
in all salt mixes. Once again, Dr. Shimek’s study indicates
lower levels for Bio-Sea MarineMix and Crystal Sea Bio-Assay than
for Instant Ocean and Coralife. In fact, Instant Ocean tested
with the second lowest Copper concentration. The other two salts
in his study were roughly in the middle of the field.

Lithium (ppm)

Lithium is a lightweight metal that exists in sea water as a
free positive ion. Lithium levels were below or in the general
range of NSW levels for all salts except those from Marine
Enterprises: Crystal Sea MarineMix and Crystal Sea Bio-Assay.
Those two salt mixes were approximately 70 times NSW levels.

Manganese (ppb)

Manganese is another potentially toxic element which was
reported in Dr. Shimek’s study to be significant higher in
Instant Ocean and Coralife than in Bio-Sea MarineMix or Crystal
Sea Bio-Assay. Our results show that, while all salt mixes were
higher than NSW, Instant Ocean was comparable to the other salts.
Only Oceanic showed a significantly high level – about ten times
the other salt mixes.

Molybdenum (ppb)

Once again, Dr. Shimek’s study showed much higher levels
in Instant Ocean and Coralife than in Bio-Sea MarineMix or
Crystal Sea Bio-Assay. However, our results show that Instant
Ocean and Bio-Sea MarineMix both have the same low level and
Crystal Sea Bio-Assay has the second highest Molybdenum
concentration.

Nickel (ppb)

Nickel is another metal that was reported in Dr. Shimek’s
study to be significant higher in Instant Ocean and Coralife than
in Bio-Sea MarineMix or Crystal Sea Bio-Assay. In fact, Crystal
Sea Bio-Assay had the highest Nickel concentration and Instant
Ocean was the third lowest.

Lead (ppb)

Lead is a toxic heavy metal that was reported in Dr.
Shimek’s study to be over 100 times higher in Instant Ocean
than in Crystal Sea Bio-Assay. Our results show that all salt
mixes were higher than NSW, but Crystal Sea Bio-Assay was in fact
slightly higher in Lead levels than Instant Ocean. Bio-Sea
MarineMix had the same level as Instant Ocean.

Palladium (ppb)

Palladium was not specifically called out in Dr. Shimek’s
study. All salt mixes were about 100,000 times higher than NSW,
but similar to each other within a factor of ten.

Selenium (ppb)

Selenium is another potential heavy metal toxin that was not
called out in Dr. Shimek’s study. The salts from Marine
Enterprises tested lower than other salts for this element.
However, all salt mixes were 100 to 400 times the average NSW
level.

Silicon (ppm)

Strontium (ppm)

Strontium is an essential element for stony coral growth.
Oceanic and Crystal Sea MarineMix showed deficient levels of
Strontium, whereas Omega Sea tested at roughly double NSW levels.
All other salt mixes were similar to NSW.

Titanium (ppb)

Titanium is yet another element that was reported in Dr.
Shimek’s study to be significantly higher in Instant Ocean
and Coralife than in Crystal Sea Bio-Assy. Our results show that
four of the salt mixes are roughly eight times NSW level:
AquaMedic, Crystal Sea MarineMix, Instant Ocean, and Crystal Sea
Bio-Assay. All other salts were two to three times NSW.

Zinc (ppb)

Zinc was also reported in Dr. Shimek’s study as being
several times higher in Instant Ocean and Coralife than Crystal
Sea Bio-Assay or Bio-Sea MarineMix. We show that Oceanic has a
significantly higher than NSW level of Zinc, and that AquaMedic,
Omega Sea and Tropic Marin are about five times the average NSW
level. Other salts are at or below NSW levels.

Summary

There are three separate areas to be addressed:

Does the manufacturer’s data hold up to close
scrutiny?

Are there heavy metal concentrations revealed in these
tests that could account for Dr. Shimek’s bio-assay
results?

Do these results shed any light on the salt mix choices
available to the hobbyist?

These items will be discussed in Part II of this article next
month.

References

Atkinson, M. and C. Bingman 1999. The Composition of
Several Synthetic Seawater

End Notes

1 “This study has demonstrated
that the artificial sea water made using some common and
popular commercial artificial salt water mixes is toxic to
sea urchin larvae using a variant of a standard bioassay.
Such water will also likely have effects on other
animals.” Shimek, 2003

2 “This study also showed that
some artificial sea water mixes produced water that could
support larval development as well as could natural
seawater.” Shimek, 2003

3 “The two salts that made
artificial seawater with the lowest survivorship of larvae
consistently have heavy metals concentrations hundreds to
hundreds of thousands times those found in natural
seawater. Those salts that had the best survival had heavy
metals concentrations that generally ranged about from, at
worst, about one third to, at best, one thousandth those
values.” Shimek, 2003

4 “â€¦ I had to rely on data
provided by the manufacturer of Crystal Sea Marinemix
Bioassay Formula. For the Bio-Sea Marinemix, I used the
data from one of the advertising brochures describing the
salt.” Shimek, 2003

5 “Fortunately, there is noa priori reason to doubt the veracity of either
of these sources.” Shimek, 2003